scholarly journals Activation of endogenous GABAA channels on airway smooth muscle potentiates isoproterenol-mediated relaxation

2008 ◽  
Vol 295 (6) ◽  
pp. L1040-L1047 ◽  
Author(s):  
George Gallos ◽  
Neil R. Gleason ◽  
Yi Zhang ◽  
Sang-Woo Pak ◽  
J. R. Sonett ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Airway Smooth Muscle ◽  
Smooth Muscles ◽  
Airway Disease ◽  
Airway Smooth Muscle Cells ◽  
Neurokinin A ◽  
Therapeutic Implications ◽  
Reactive Airway Disease ◽  
Endogenous Gaba

Reactive airway disease predisposes patients to episodes of acute smooth muscle mediated bronchoconstriction. We have for the first time recently demonstrated the expression and function of endogenous ionotropic GABAA channels on airway smooth muscle cells. We questioned whether endogenous GABAA channels on airway smooth muscle could augment β-agonist-mediated relaxation. Guinea pig tracheal rings or human bronchial airway smooth muscles were equilibrated in organ baths with continuous digital tension recordings. After pretreatment with or without the selective GABAA antagonist gabazine (100 μM), airway muscle was contracted with acetylcholine or β-ala neurokinin A, followed by relaxation induced by cumulatively increasing concentrations of isoproterenol (1 nM to 1 μM) in the absence or presence of the selective GABAA agonist muscimol (10–100 μM). In separate experiments, guinea pig tracheal rings were pretreated with the large conductance KCa channel blocker iberiotoxin (100 nM) after an EC50 contraction with acetylcholine but before cumulatively increasing concentrations of isoproterenol (1 nM to 1 uM) in the absence or presence of muscimol (100 uM). GABAA activation potentiated the relaxant effects of isoproterenol after an acetylcholine or tachykinin-induced contraction in guinea pig tracheal rings or an acetylcholine-induced contraction in human endobronchial smooth muscle. This muscimol-induced potentiation of relaxation was abolished by gabazine pretreatment but persisted after blockade of the maxi KCa channel. Selective activation of endogenous GABAA receptors significantly augments β-agonist-mediated relaxation of guinea pig and human airway smooth muscle, which may have important therapeutic implications for patients in severe bronchospasm.

scholarly journals Propofol Preferentially Relaxes Neurokinin Receptor-2-induced Airway Smooth Muscle Contraction in Guinea Pig Trachea

2010 ◽  
Vol 112 (6) ◽  
pp. 1335-1344 ◽  
Author(s):  
Neil R. Gleason ◽  
George Gallos ◽  
Yi Zhang ◽  
Charles W. Emala
Keyword(s):  
Smooth Muscle ◽  
Substance P ◽  
Guinea Pig ◽  
Airway Smooth Muscle ◽  
Electrical Field Stimulation ◽  
Airway Disease ◽  
Smooth Muscle Contraction ◽  
Neurokinin A ◽  
Protective Effects ◽  
Reactive Airway Disease

Background Propofol is the anesthetic of choice for patients with reactive airway disease and is thought to reduce intubation- or irritant-induced bronchoconstriction by decreasing the cholinergic component of vagal nerve activation. However, additional neurotransmitters, including neurokinins, play a role in irritant-induced bronchoconstriction. We questioned the mechanistic assumption that the clinically recognized protective effect of propofol against irritant-induced bronchoconstriction during intubation was due to attenuation of airway cholinergic reflexes. Methods Muscle force was continuously recorded from isolated guinea pig tracheal rings in organ baths. Rings were subjected to exogenous contractile agonists (acetylcholine, histamine, endothelin-1, substance P, acetyl-substance P, and neurokinin A) or to electrical field stimulation (EFS) to differentiate cholinergic or nonadrenergic, noncholinergic nerve-mediated contraction with or without cumulatively increasing concentrations of propofol, thiopental, etomidate, or ketamine. Results Propofol did not attenuate the cholinergic component of EFS-induced contraction at clinically relevant concentrations. In contrast, propofol relaxed nonadrenergic, noncholinergic-mediated EFS contraction at concentrations within the clinical range (20-100 mum, n = 9; P < 0.05), and propofol was more potent against an exogenous selective neurokinin-2 receptor versus neurokinin-1 receptor agonist contraction (n = 6, P < 0.001). Conclusions Propofol, at clinically relevant concentrations, relaxes airway smooth muscle contracted by nonadrenergic, noncholinergic-mediated EFS and exogenous neurokinins but not contractions elicited by the cholinergic component of EFS. These findings suggest that the mechanism of protective effects of propofol against irritant-induced bronchoconstriction involves attenuation of tachykinins released from nonadrenergic, noncholinergic nerves acting at neurokinin-2 receptors on airway smooth muscle.

scholarly journals Airway epithelium is a predominant source of endogenous airway GABA and contributes to relaxation of airway smooth muscle tone

2013 ◽  
Vol 304 (3) ◽  
pp. L191-L197 ◽  
Author(s):  
George Gallos ◽  
Elizabeth Townsend ◽  
Peter Yim ◽  
Laszlo Virag ◽  
Yi Zhang ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Airway Smooth Muscle ◽  
Airway Epithelium ◽  
Muscle Force ◽  
Airway Smooth Muscle Cells ◽  
Human Airway ◽  
Gaba Transporters ◽  
Endogenous Gaba ◽  
Cellular Source

Chronic obstructive pulmonary disease and asthma are characterized by hyperreactive airway responses that predispose patients to episodes of acute airway constriction. Recent studies suggest a complex paradigm of GABAergic signaling in airways that involves GABA-mediated relaxation of airway smooth muscle. However, the cellular source of airway GABA and mechanisms regulating its release remain unknown. We questioned whether epithelium is a major source of GABA in the airway and whether the absence of epithelium-derived GABA contributes to greater airway smooth muscle force. Messenger RNA encoding glutamic acid decarboxylase (GAD) 65/67 was quantitatively measured in human airway epithelium and smooth muscle. HPLC quantified GABA levels in guinea pig tracheal ring segments under basal or stimulated conditions with or without epithelium. The role of endogenous GABA in the maintenance of an acetylcholine contraction in human airway and guinea pig airway smooth muscle was assessed in organ baths. A 37.5-fold greater amount of mRNA encoding GAD 67 was detected in human epithelium vs. airway smooth muscle cells. HPLC confirmed that guinea pig airways with intact epithelium have a higher constitutive elution of GABA under basal or KCl-depolarized conditions compared with epithelium-denuded airway rings. Inhibition of GABA transporters significantly suppressed KCl-mediated release of GABA from epithelium-intact airways, but tetrodotoxin was without effect. The presence of intact epithelium had a significant GABAergic-mediated prorelaxant effect on the maintenance of contractile tone. Airway epithelium is a predominant cellular source of endogenous GABA in the airway and contributes significant prorelaxant GABA effects on airway smooth muscle force.

IL-1 beta and IL-6 induce hyperplasia and hypertrophy of cultured guinea pig airway smooth muscle cells

1995 ◽  
Vol 78 (4) ◽  
pp. 1555-1563 ◽  
Author(s):  
S. De ◽  
E. T. Zelazny ◽  
J. F. Souhrada ◽  
M. Souhrada
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Protein Content ◽  
Airway Smooth Muscle ◽  
Dna Content ◽  
Thymidine Incorporation ◽  
Interleukin 1 ◽  
Total Protein Content ◽  
Leucine Incorporation ◽  
Airway Smooth Muscle Cells

Guinea pig airway smooth muscle (ASM) cells were maintained in a primary tissue culture (passages 1–3). Cells were exposed to human recombinant interleukin-1 beta (IL-1 beta; 20–100 pg/ml) or interleukin-6 (IL-6; 1–4 ng/ml) in the presence of indomethacin (1 microgram/ml) for up to 5 days. Proliferation of ASM cells was assessed with two techniques, direct counting of cells with a hemacytometer and [3H]thymidine incorporation corrected for total protein content. Hypertrophy of ASM cells was assessed by [3H]leucine incorporation (evaluation of protein synthesis), determination of total DNA content, DNA content per cell, and protein content per cell. We observed that the exposure of ASM cells to human recombinant IL-1 beta or IL-6, in all studied concentrations, significantly increased the number of cells as well as [3H]thymidine incorporation into ASM cells. We also found that exposure of ASM to these two cytokines increased [3H]leucine incorporation into the ASM cells and increased protein content and DNA content per single cell. These changes were also concentration dependent. We conclude that the two proinflammatory cytokines, IL-1 beta and IL-6, which are present in asthmatic lungs, increased the proliferation of ASM cells (hyperplasia) as well as their overall size and size of their nuclei, as measured by biochemical markers. These findings are compatible with the presence of ASM hypertrophy.

Coupling mechanisms in airway smooth muscle

1990 ◽  
Vol 258 (4) ◽  
pp. L119-L133 ◽  
Author(s):  
R. F. Coburn ◽  
C. B. Baron
Keyword(s):  
Smooth Muscle ◽  
Membrane Potential ◽  
Airway Smooth Muscle ◽  
Electromechanical Coupling ◽  
Cell Function ◽  
Surface Membrane ◽  
Smooth Muscles ◽  
Membrane Receptors ◽  
Airway Smooth Muscle Cells ◽  
Coupling Mechanisms

This review documents available information about coupling mechanisms involved in airway smooth muscle force development and maintenance and relaxation of force. Basic concepts, obtained from experiments performed on many different mammalian cell types, are in place regarding coupling between surface membrane receptors and cell function; these concepts are considered as a framework for understanding coupling between receptors and contractile proteins in smooth muscles and in airway smooth muscles. We have divided various components of coupling mechanisms into those dependent on changes in the surface membrane potential (electromechanical coupling) and those independent of the surface membrane potential (pharmacomechanical coupling). We have, to some degree, emphasized modulation of coupling mechanisms by intrasurface membrane microprocessing or by second messengers. A challenge for the future is to obtain a better understanding of how coupling mechanisms are altered or modulated during different phases of contractions evoked by a single agonist and under conditions of multiple agonist exposure to airway smooth muscle cells.

scholarly journals Specific detection of the endogenous transient receptor potential (TRP)-1 protein in liver and airway smooth muscle cells using immunoprecipitation and Western-blot analysis

2002 ◽  
Vol 364 (3) ◽  
pp. 641-648 ◽  
Author(s):  
Hwei Ling ONG ◽  
Jinglong CHEN ◽  
Tim CHATAWAY ◽  
Helen BRERETON ◽  
Lei ZHANG ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Guinea Pig ◽  
Western Blot ◽  
Airway Smooth Muscle ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Airway Smooth Muscle Cells ◽  
Transient Receptor

Although there are numerous reports of the presence of mRNA encoding the transient receptor potential (TRP)-1 protein in animal cells and of the detection of the heterologously expressed TRP-1 protein by Western-blot analysis, it has proved difficult to unequivocally detect endogenous TRP-1 proteins. A combination of immunoprecipitation and Western-blot techniques, employing a polyclonal antibody and a monoclonal antibody respectively, was developed. Using this technique, a band of approx. 80kDa was detected in extracts of H4-IIE rat liver hepatoma cell line and guinea-pig airway smooth muscle (ASM) cells transfected with human TRPC-1 cDNA. In extracts of untransfected H4-IIE cells, ASM cells, rat brain and guinea-pig brain, a band of approx. 92kDa was detected. Reverse transcriptase PCR experiments detected cDNA encoding both the α- and β-isoforms of TRP-1 in H4-IIE cells. Treatment of protein extracts with peptide N-glycosidase F indicated that the 92kDa band represents an N-glycosylated protein. Western blots conducted with a commercial polyclonal anti-(TRP-1) antibody (Alm) detected a band of 120kDa in extracts of H4-IIE cells and guinea-pig ASM cells. A combination of immunoprecipitation and Western-blotting techniques with the Alm antibody did not detect any bands at 92kDa or 120kDa in extracts of H4-IIE and ASM cells. It is concluded that (a) the 92-kDa band detected in untransfected H4-IIE and ASM cells corresponds to the N-glycosylated β-isoform of endogenous TRP-1, (b) the combined immunoprecipitation and Western-blot approach, employing two different antibodies, provides a reliable and specific procedure for detecting endogenous TRP-1 proteins, and (c) that caution is required in developing and utilizing anti-(TRP-1) antibodies.

Substance P activates Cl− and K+ conductances in guinea-pig tracheal smooth muscle cells

1994 ◽  
Vol 72 (6) ◽  
pp. 705-710 ◽  
Author(s):  
Luke J. Janssen ◽  
Stephen M. Sims
Keyword(s):  
Smooth Muscle ◽  
Substance P ◽  
Guinea Pig ◽  
Smooth Muscle Cells ◽  
Voltage Clamp ◽  
Airway Smooth Muscle ◽  
Muscle Cells ◽  
Equilibrium Potential ◽  
Airway Smooth Muscle Cells ◽  
Inward Current

Substance P (SP) causes bronchoconstriction, but its effects on airway smooth muscle ion conductances are unknown. We investigated the effects of SP on single smooth muscle cells dissociated from guinea-pig trachealis. Under voltage clamp at −60 mV, SP evoked reversible contractions and inward current (ISP). ISP had a latency of approximately 1 s, reached a peak of 1039 ± 147 pA (n = 19) about 2 s after onset of application, and declined to baseline levels over the next 5–10 s. At more positive holding potentials (−25 and 0 mV), the inward current was decreased in magnitude and preceded by outward current. With 140 mM K+ in the electrode and Cl− equilibrium potential (ECl) of about 0 mV, ISP was outwardly rectifying and reversed at −11 ± 2 mV. When K+ currents were blocked using Cs+, the current–voltage relationship for ISP was linear and reversed at 3 ± 1 mV. The reversal potential was dependent on the Cl− gradient across the membrane. These results suggest that SP caused a transient activation of Cl− and K+ conductances. Following the initial transient inward current, SP caused a prolonged suppression of spontaneously active K+ currents. The findings that SP evoked contractions during voltage clamp at potentials at which voltage-dependent Ca2+ channels are not active, and that current oscillations were also evoked by SP, suggest that SP is acting through release of Ca2+ from internal stores. Furthermore, SP occluded the inward current evoked by acetylcholine, suggesting that the peptidergic and cholinergic signalling pathways converge. We conclude that SP releases Ca2+ from internal stores in guinea-pig airway smooth muscle cells, leading to activation of Cl− and K+ conductances, depolarization, and contraction.Key words: Ca2+-dependent conductances, spontaneous transient outward currents, acetylcholine.

A method for isolating adult and neonatal airway smooth muscle cells and measuring shortening velocity

1999 ◽  
Vol 86 (1) ◽  
pp. 427-435 ◽  
Author(s):  
S. P. Driska ◽  
R. E. Laudadio ◽  
M. R. Wolfson ◽  
T. H. Shaffer
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Single Cell ◽  
Airway Smooth Muscle ◽  
Smooth Muscles ◽  
Muscle Cells ◽  
Cell Length ◽  
Airway Smooth Muscle Cells ◽  
Shortening Velocity ◽  
Adult Cells

Methods are described for isolating smooth muscle cells from the tracheae of adult and neonatal sheep and measuring the single-cell shortening velocity. Isolated cells were elongated, Ca2+ tolerant, and contracted rapidly and substantially when exposed to cholinergic agonists, KCl, serotonin, or caffeine. Adult cells were longer and wider than preterm cells. Mean cell length in 1.6 mM CaCl2 was 194 ± 57 (SD) μm ( n = 66) for adult cells and 93 ± 32 μm ( n = 20) for preterm cells ( P < 0.05). Mean cell width at the widest point of the adult cells was 8.2 ± 1.8 μm ( n = 66) and 5.2 ± 1.5 μm ( n = 20) for preterm cells ( P < 0.05). Cells were loaded into a perfusion dish maintained at 35°C and exposed to agonists, and contractions were videotaped. Cell lengths were measured from 30 video frames and plotted as a function of time. Nonlinear fitting of cell length to an exponential model gave shortening velocities faster than most of those reported for airway smooth muscle tissues. For a sample of 10 adult and 10 preterm cells stimulated with 100 μM carbachol, mean (± SD) shortening velocity of the preterm cells was not different from that of the adult cells (0.64 ± 0.30 vs. 0.54 ± 0.27 s−1, respectively), but preterm cells shortened more than adult cells (68 ± 12 vs. 55 ± 11% of starting length, respectively; P < 0.05). The preparative and analytic methods described here are widely applicable to other smooth muscles and will allow contraction to be studied quantitatively at the single-cell level.

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